Extraction and fractionation of cocoa proteins as applied to several varieties of cocoa beans

Zak, Dennis L.; Keeney, P. G.

doi:10.1021/jf60205a055

Cited by 28 publications

(27 citation statements)

References 12 publications

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“…The polypeptide bands were similar to those obtained by several researchers for cacao beans in classical fermentation [26,37,38]. Glutelin fraction from UF cacao had proteins with molecular mass > 200 kDa and others of low molecular weight, 19.7 and 14.4 kDa, while the SFD cacao fraction presented only high molecular mass proteins (> 200 kDa) that were slightly higher than those reported previously [39]. …”

Section: Resultssupporting

confidence: 86%

Antitumor activity against murine lymphoma L5178Y model of proteins from cacao (Theobroma cacao L.) seeds in relation with in vitro antioxidant activity

Preza

Jaramillo

Puebla

et al. 2010

BMC Complement Altern Med

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BackgroundRecently, proteins and peptides have become an added value to foodstuffs due to new knowledge about its structural analyses as related to antioxidant and anticancer activity. Our goal was to evaluate if protein fractions from cacao seeds show antitumor activity on lymphoma murine L5178Y model. The antioxidant activity of these fractions was also evaluated with the aim of finding a correlation with the antitumor activity.MethodsDifferential extraction of proteins from unfermented and semi-fermented-dry cacao seeds was performed and characterized by SDS-PAGE and FPLC size-exclusion chromatography. Antitumor activity was evaluated against murine lymphoma L5178Y in BALB/c mice (6 × 104 cells i.p.), with a treatment oral dose of 25 mg/kg/day of each protein fraction, over a period of 15 days. Antioxidant activity was evaluated by the ABTS+ and ORAC-FL assays.ResultsAlbumin, globulin and glutelin fractions from both cacao seed type were obtained by differential solubility extraction. Glutelins were the predominant fraction. In the albumin fraction, polypeptides of 42.3 and 8.5 kDa were found in native conditions, presumably in the form of two peptide chains of 21.5 kDa each one. The globulin fraction presented polypeptides of 86 and 57 kDa in unfermented cacao seed that produced the specific-cacao aroma precursors, and after fermentation the polypeptides were of 45 and 39 kDa. The glutelin fraction presented proteins >200 kDa and globulins components <100 KDa in lesser proportion. Regarding the semifermented-dry cacao seed, it was observed that the albumin fraction showed antitumoral activity, since it caused significant decreases (p < 0.05) in the ascetic fluid volume and packed cell volume, inhibiting cell growth in 59.98 ± 13.6% at 60% of the population; while the greatest antioxidant capacity due to free radical scavenging capacity was showed by the albumin and glutelin fraction in both methods assayed.ConclusionThis study is the first report on the biological activity of semifermented-dry cacao protein fractions with their identification, supporting the traditional use of the plant. The albumin fraction showed antitumor and free radical scavenging capacity, however both activities were not correlated. The protein fractions could be considered as source of potential antitumor peptides.

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Section: Resultssupporting

confidence: 86%

Antitumor activity against murine lymphoma L5178Y model of proteins from cacao (Theobroma cacao L.) seeds in relation with in vitro antioxidant activity

Preza

Jaramillo

Puebla

et al. 2010

BMC Complement Altern Med

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“…However, cacao has received little attention with respect to proteomic research and until now, no study has been carried out to analyse the protein pattern of cacao during its infection by pests or diseases. Some studies to understand and improve the chocolate quality and flavor by characterization of protein changes during ripening of the cacao fruit, postharvest fermentation of cacao beans and key processing steps in the chocolate factory, namely, roasting and conching, were developed [10,11]. However, these studies carried out for cacao beans, were not related to the high level of polysaccharides present in the vegetative organs such as leaves and meristems [12,13], which are specifically infected by the hemibiotrophic fungus M. perniciosa.…”

Section: Introductionmentioning

confidence: 99%

Protein extraction for proteome analysis from cacao leaves and meristems, organs infected by Moniliophthora perniciosa, the causal agent of the witches' broom disease

Pirovani

Carvalho

Machado³

et al. 2008

Electrophoresis

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Preparation of high-quality proteins from cacao vegetative organs is difficult due to very high endogenous levels of polysaccharides and polyphenols. In order to establish a routine procedure for the application of proteomic and biochemical analysis to cacao tissues, three new protocols were developed; one for apoplastic washing fluid (AWF) extraction, and two for protein extraction--under denaturing and nondenaturing conditions. The first described method allows a quick and easy collection of AWF--using infiltration-centrifugation procedure--that is representative of its composition in intact leaves according to the smaller symplastic contamination detected by the use of the hexose phosphate isomerase marker. Protein extraction under denaturing conditions for 2-DE was remarkably improved by the combination of chemically and physically modified processes including phenol, SDS dense buffer and sonication steps. With this protocol, high-quality proteins from cacao leaves and meristems were isolated, and for the first time well-resolved 1-DE and 2-DE protein patterns of cacao vegetative organs are shown. It also appears that sonication associated with polysaccharide precipitation using tert-butanol was a crucial step for the nondenaturing protein extraction and subsequent enzymatic activity detection. It is expected that the protocols described here could help to develop high-level proteomic and biochemical studies in cacao also being applicable to other recalcitrant plant tissues.

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“…This aspect cannot be ascribed to instrumental factors, but to the ineffectiveness of our sample treatment procedure to extract this protein. Indeed, Zak et al have shown that vicilin is hydrosoluble only in presence of high salt concentrations. During preparation of the samples for MALDI analysis, desalification of the powder has been performed by dialysis and, after dialysis and treatment with ultrasonic wave, the supernatant has been collected.…”

Section: Resultsmentioning

confidence: 99%

“…In particular, cocoa proteins (representing 10–15% of dry bean weight) affect the characteristics of chocolate. The four predominant protein fractions, representing 95% (w/w) of total cocoa seed proteins, are albumins (water‐soluble), globulins (salt‐soluble), prolamins (alcohol‐soluble) and glutelins (soluble in dilute acids and alkali) . Voigt et al found that the protein content of cocoa beans is composed of 52% and 43% of albumin and globulin fractions, respectively.…”

mentioning

confidence: 99%

The protein profile of Theobroma cacao L. seeds as obtained by matrix‐assisted laser desorption/ionization mass spectrometry

Bertazzo

Agnolin

Comai

et al. 2011

Rapid Comm Mass Spectrometry

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The water-soluble protein profile of the seeds of green, red, and yellow Theobroma cacao L. fruits has been determined by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-ToF-MS). The seeds were powdered under liquid nitrogen and defatted. The residues were dialyzed and lyophilized. The obtained samples were suspended in the matrix solution of sinapinic acid. The obtained MALDI mass spectra showed the presence of a wide number of proteins with molecular weight ranging from 8000 to 13,000 Da and a cluster of peaks centered at 21,000 Da that were attributed to albumin. The abundance of this peak was found to depend on the different portion of the seed (husk, apical and cortical parts); however, the MALDI mass spectra obtained from the different varieties of cocoa were practically superimposable. Changes in the protein profiles were also observed after the cocoa seeds were treated by fermentation and roasting, which are processes usually employed for the commercial production of cocoa.

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Extraction and fractionation of cocoa proteins as applied to several varieties of cocoa beans

Cited by 28 publications

References 12 publications

Antitumor activity against murine lymphoma L5178Y model of proteins from cacao (Theobroma cacao L.) seeds in relation with in vitro antioxidant activity

Antitumor activity against murine lymphoma L5178Y model of proteins from cacao (Theobroma cacao L.) seeds in relation with in vitro antioxidant activity

Protein extraction for proteome analysis from cacao leaves and meristems, organs infected by Moniliophthora perniciosa, the causal agent of the witches' broom disease

The protein profile of Theobroma cacao L. seeds as obtained by matrix‐assisted laser desorption/ionization mass spectrometry

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